CRISPR/Cas9-mediated <i>ELANE</i> knockout enables neutrophilic maturation of primary hematopoietic stem and progenitor cells and induced pluripotent stem cells of severe congenital neutropenia patients

Nasri, Masoud; Ritter, Malte; Mir, Perihan; Dannenmann, Benjamin; Aghaallaei, Narges; Amend, Diana; Makaryan, Vahagn; Xu, Yitong; Fletcher, Breanna; Bernhard, Regine; Steiert, I.; Hähnel, Karin; Berger, Jürgen; Koch, Iris; Sailer, Brigitte; Hipp, Katharina; Zeidler, Cornelia; Klimiankou, Maksim; Bajoghli, Baubak; Dale, David C.; Welte, Karl; Skokowa, Julia

doi:10.3324/haematol.2019.221804

Cited by 42 publications

(43 citation statements)

References 44 publications

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“…Recent studies have shown that ELANE whole gene deletion mutation does not cause neutropenia in humans and mice ( 45 ). Interestingly, ELANE knockout restores the granulopoiesis and increases differentiation into functional mature neutrophils in vitro ( 47 ). Therefore, we expect the neutrophil elastase gene to be a potential gene therapy target in patients with ELANE mutations, especially those who do not respond to G-CSF or who are at high risk of malignant transformation.…”

Section: Discussionmentioning

confidence: 99%

“…Interestingly, humans ( 45 ) and mice ( 46 ) with ELANE biallelic loss of function defect produce functional neutrophils with normal complete blood counts. Moreover, knock-out of the mutant allele in hematopoietic stem cells derived from SCN patients restores neutrophils maturation ( 47 ). This excludes the NE haploinsufficiency as a pathomechanism of SCN, and it is likely that the cause of neutropenia is not the lack of neutrophil elastase itself, but protease malfunction.…”

Section: Elane Mutations In Pathogenesis Of Congenital Neutropeniamentioning

confidence: 99%

“…They observed granulocytic differentiation as well as phagocytic ability, reactive oxygen species production and chemotaxis by neutrophils without ELANE in SCN patient-derived iPSc, primary hematopoietic stem cells and HL-60 cell line with ELANE KO. It suggests an alternative method of SCN treatment using ex vivo CRISPR/Cas9 ribonucleoprotein mediated ELANE KO ( 47 ). An alternated approach to repair ELANE mutations was performed using two different single guide RNA (sgRNA) targeting both mutant allele and exon 4.…”

Section: Clinical Features and Therapy Of Scn With Elane Mutationsmentioning

confidence: 99%

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Neutrophil Elastase Defects in Congenital Neutropenia

et al. 2021

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Severe congenital neutropenia (SCN) is a rare hematological condition with heterogenous genetic background. Neutrophil elastase (NE) encoded by ELANE gene is mutated in over half of the SCN cases. The role of NE defects in myelocytes maturation arrest in bone marrow is widely investigated; however, the mechanism underlying this phenomenon has still remained unclear. In this review, we sum up the studies exploring mechanisms of neutrophil deficiency, biological role of NE in neutrophil and the effects of ELANE mutation and neutropenia pathogenesis. We also explain the hypotheses presented so far and summarize options of neutropenia therapy.

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Section: Discussionmentioning

confidence: 99%

Section: Elane Mutations In Pathogenesis Of Congenital Neutropeniamentioning

confidence: 99%

Section: Clinical Features and Therapy Of Scn With Elane Mutationsmentioning

confidence: 99%

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Neutrophil Elastase Defects in Congenital Neutropenia

et al. 2021

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“…In this respect, the infusion of ex vivo expanded neutrophils to overcome neutropenia, as has e.g., been explored in early clinical trials using neutrophils expanded from CD34 + hematopoietic stem cells [ 252 , 253 ], may well be combined with immunomodulatory strategies that ensure an anticancer polarization of neutrophils, such as CRISPR/Cas9 gene editing of N2-polarizing transcription factors. Such gene editing of hematopoietic stem cells can be achieved with high efficiency [ 254 ], with proof-of-concept for the effective modification of neutrophil activity recently generated for severe congenital neutropenia using ELANE knock-out [ 255 ] that in phagocytic functions, ROS production, and chemotaxis was similar to healthy donors.…”

Section: Discussionmentioning

confidence: 99%

The Neutrophil: The Underdog That Packs a Punch in the Fight against Cancer

Avtenyuk

Visser

Bremer

et al. 2020

IJMS

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The advent of immunotherapy has had a major impact on the outcome and overall survival in many types of cancer. Current immunotherapeutic strategies typically aim to (re)activate anticancer T cell immunity, although the targeting of macrophage-mediated anticancer innate immunity has also emerged in recent years. Neutrophils, although comprising ≈ 60% of all white blood cells in the circulation, are still largely overlooked in this respect. Nevertheless, neutrophils have evident anticancer activity and can induce phagocytosis, trogocytosis, as well as the direct cytotoxic elimination of cancer cells. Furthermore, therapeutic tumor-targeting monoclonal antibodies trigger anticancer immune responses through all innate Fc-receptor expressing cells, including neutrophils. Indeed, the depletion of neutrophils strongly reduced the efficacy of monoclonal antibody treatment and increased tumor progression in various preclinical studies. In addition, the infusion of neutrophils in murine cancer models reduced tumor progression. However, evidence on the anticancer effects of neutrophils is fragmentary and mostly obtained in in vitro assays or murine models with reports on anticancer neutrophil activity in humans lagging behind. In this review, we aim to give an overview of the available knowledge of anticancer activity by neutrophils. Furthermore, we will describe strategies being explored for the therapeutic activation of anticancer neutrophil activity.

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“…Gene editing approach based on CRISPR/Cas9-sgRNA has also been recently shownto reduce ELANE expression ex vivo in bone marrow hematopoietic progenitors from patients with SCN. CRISPR-Cas9-mediated knockdown of ELANE significantly induced neutrophil maturation in vitro [244]. It should be nevertheless noted that CRISPR-Cas9 technology presents several limitations for its rapid translation as a therapy, including cell type dependent delivery, incomplete efficiency of homologous recombination and the possibility of off-target editing [245].…”

Section: Discussion and Perspectivesmentioning

confidence: 99%

Nonsense Suppression Therapy: New Hypothesis for the Treatment of Inherited Bone Marrow Failure Syndromes

Bezzerri

Api

Allegri

et al. 2020

IJMS

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Inherited bone marrow failure syndromes (IBMFS) are a group of cancer-prone genetic diseases characterized by hypocellular bone marrow with impairment in one or more hematopoietic lineages. The pathogenesis of IBMFS involves mutations in several genes which encode for proteins involved in DNA repair, telomere biology and ribosome biogenesis. The classical IBMFS include Shwachman–Diamond syndrome (SDS), Diamond–Blackfan anemia (DBA), Fanconi anemia (FA), dyskeratosis congenita (DC), and severe congenital neutropenia (SCN). IBMFS are associated with high risk of myelodysplastic syndrome (MDS), acute myeloid leukemia (AML), and solid tumors. Unfortunately, no specific pharmacological therapies have been highly effective for IBMFS. Hematopoietic stem cell transplantation provides a cure for aplastic or myeloid neoplastic complications. However, it does not affect the risk of solid tumors. Since approximately 28% of FA, 24% of SCN, 21% of DBA, 20% of SDS, and 17% of DC patients harbor nonsense mutations in the respective IBMFS-related genes, we discuss the use of the nonsense suppression therapy in these diseases. We recently described the beneficial effect of ataluren, a nonsense suppressor drug, in SDS bone marrow hematopoietic cells ex vivo. A similar approach could be therefore designed for treating other IBMFS. In this review we explain in detail the new generation of nonsense suppressor molecules and their mechanistic roles. Furthermore, we will discuss strengths and limitations of these molecules which are emerging from preclinical and clinical studies. Finally we discuss the state-of-the-art of preclinical and clinical therapeutic studies carried out for IBMFS.

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CRISPR/Cas9-mediated ELANE knockout enables neutrophilic maturation of primary hematopoietic stem and progenitor cells and induced pluripotent stem cells of severe congenital neutropenia patients

Cited by 42 publications

References 44 publications

Neutrophil Elastase Defects in Congenital Neutropenia

Neutrophil Elastase Defects in Congenital Neutropenia

The Neutrophil: The Underdog That Packs a Punch in the Fight against Cancer

Nonsense Suppression Therapy: New Hypothesis for the Treatment of Inherited Bone Marrow Failure Syndromes

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